Although this invention is not limited to cuvettes used for nucleic acid amplification, the background is described in the context of the latter, as such amplification led to the invention.
Nucleic acid amplification generally proceeds via a particular protocol. One useful protocol is that set forth in U.S. Pat. No. 4,683,195. Briefly, that protocol features, in the case of DNA amplification, the following:
(1) A complete DNA double helix is optionally chemically excised, using an appropriate restriction enzyme(s), to isolate the region of interest. PA0 (2) A solution of the isolated nucleic acid portion (here, DNA) and nucleotides is heated to and maintained at 92.degree.-95.degree. C. for a length of time, e.g., no more than about 10 minutes, to denature the two nucleic acid strands; i.e., cause them to unwind and separate and form a template. PA0 (3) The solution is then cooled through a 50.degree.-60.degree. C. zone to cause a primer nucleic acid strand to anneal or "attach" to each of the two template strands. To make sure this happens, the solution is held at an appropriate temperature, such as about 55.degree. C. for about 15 seconds, in an "incubation" zone. PA0 (4) The solution is then heated to and held at about 70.degree. C., to cause an extension enzyme, preferably a thermostatable enzyme, to extend the primer strand bound to the template strand by using the nucleotides that are present. PA0 (5) The completed new pair of strands is heated to 92.degree.-95.degree. C. again, for about 10-15 seconds, to cause this pair to separate. PA0 (6) Steps (3)-(5) are then repeated, a number of times until the appropriate number of strands are obtained. The more repetitions, the greater the number of multiples of the nucleic acid (here, DNA) that is produced. Preferably the desired concentration of nucleic acid is reached in a minimum amount of time.
A cuvette is usually used to hold the solution while it passes through the aforementioned temperature stages. Depending upon the design given to the cuvette, it can proceed more or less rapidly through the various stages. A key aspect controlling this is the thermal transfer efficiency of the cuvette--that is, its ability to transfer heat more or less instantaneously to or from all of the liquid solution within the cuvette. The disposition and the thermal resistance of the liquid solution itself are usually the major aspects affecting the thermal transfer, since portions of the liquid solution that are relatively far removed from the heat source or sink, will take longer to reach the desired temperature.
The crudest and earliest type of cuvette used in the prior art is a test tube, which has poor thermal transfer efficiency since (a) the walls of the cuvette by being glass or plastic, do not transfer thermal energy well, and (b) a cylinder of liquid has relatively poor thermal transfer throughout the liquid. That is, not only does the liquid have low thermal conductivity, but also a cylinder of liquid has a low surface to volume ratio, that is, about 27 in.sup.-1 for a fill of about 100 .mu.l.
Still another problem in DNA amplification is the manner in which the cuvette alows for ready removal of the liquid after reaction is complete. A test tube configuration readily permits such removal. However, modification of the cuvette to provide better thermal transfer efficiency tends to reduce the liquid transferability. That is, a cuvette having capillary spacing only, permits rapid heating of the contents. However, the capillary spacing resists liquid removal.